Sugihara T, Kaul S C, Mitsui Y, Wadhwa R
Institute of Applied Biochemistry, University of Tsukuba, Japan.
Biochim Biophys Acta. 1994 Dec 30;1224(3):365-70. doi: 10.1016/0167-4889(94)90269-0.
The mechanism(s) involved in immortalization that constitute the first step during malignant transformation has been the subject of our interest. By the use of spontaneously immortalized mouse embryonic fibroblasts we have earlier identified two stages of immortalization which are characterized by growth characteristics of the cells, their conditioned medium and the protein markers such as p53, p81 and mortalin (Kaul et al. (1994) Biochim. Biophys. Acta, in press). The present study was planned to purify the mitogenic factors from the conditioned medium of stage II cells. Sequential purification by chromatography followed by peptide sequencing has characterized one of these as vascular endothelial growth factor (VEGF). Further analysis by RT-PCR suggests that the spontaneously immortalized stage II fibroblasts have enhanced synthesis and secretion of VEGF as compared to their mortal parent cells. Expression of a novel 304 bp long form of VEGF is identified in immortal fibroblasts in addition to the three known alternatively spliced forms. The study points to the involvement of VEGF function during spontaneous immortalization of mouse embryonic fibroblasts.
构成恶性转化第一步的永生化所涉及的机制一直是我们感兴趣的课题。通过使用自发永生化的小鼠胚胎成纤维细胞，我们 earlier 鉴定出了永生化的两个阶段，其特征在于细胞的生长特性、它们的条件培养基以及诸如 p53、p81 和 mortalin 等蛋白质标志物（考尔等人（1994 年），《生物化学与生物物理学学报》，即将发表）。本研究旨在从 II 期细胞的条件培养基中纯化促有丝分裂因子。通过色谱法进行连续纯化，随后进行肽测序，已将其中一种鉴定为血管内皮生长因子（VEGF）。通过 RT-PCR 进一步分析表明，与它们的有限寿命亲代细胞相比，自发永生化的 II 期成纤维细胞具有增强的 VEGF 合成和分泌。除了三种已知的可变剪接形式外，在永生化成纤维细胞中还鉴定出了一种新的 304 bp 长形式的 VEGF。该研究指出了 VEGF 功能在小鼠胚胎成纤维细胞自发永生化过程中的参与。
